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Multiple research studies have demonstrated the efficacy of lactic acid bacteria in boosting both innate and adaptive immune responses. We have created a Lactococcus lactis variant that produces a modified combination protein with Fms-like tyrosine kinase 3 ligand and co-stimulator O × 40 ligand, known as HuFOLactis. The genetically modified variant was purposely created to activate T cells, NK cells, and DC cells in a laboratory setting. Furthermore, we explored the possibility of using the tumor-penetrating peptide iRGD to deliver HuFOLactis-activated immune cells to hard-to-reach tumor areas. Following brief stimulation with HuFOLactis, immune cell phenotypes and functions were assessed using flow cytometry. Confocal microscopy was employed to demonstrate the infiltrative and cytotoxic capabilities of iRGD-modified HuFOLactis-activated immune cells within tumor spheroids. The efficacy of iRGD modified HuFOLactis-activated immune cells against tumors was assessed in xenograft mouse models. HuFOLactis treatment resulted in notable immune cell activation, demonstrated by elevated levels of CD25, CD69, and CD137. Additionally, these activated immune cells showed heightened cytokine production and enhanced cytotoxicity against MKN45 cell lines. Incorporation of the iRGD modification facilitated the infiltration of HuFOLactis-activated immune cells into multicellular spheroids (MCSs). Additionally, immune cells activated by HuFOLactis and modified with iRGD, in combination with anti-PD-1 treatment, effectively halted tumor growth and prolonged survival in a mouse model of gastric cancer.

BACKGROUND: Natural killer (NK) cell-based immunotherapy is a promising treatment approach for multiple myeloma (MM), but obtaining a sufficient number of activated NK cells remains challenging. Here, we report an improved method to generate ex vivo expanded NK (eNK) cells from MM patients based on genetic engineering of K562 cells to express OX40 ligand and membrane-bound (mb) IL-18 and IL-21. METHODS: K562-OX40L-mbIL-18/-21 cells were generated by transducing K562-OX40L cells with a lentiviral vector encoding mbIL-18 and mbIL-21, and these were used as feeder cells to expand NK cells from peripheral blood mononuclear cells of healthy donors (HDs) and MM patients in the presence of IL-2/IL-15. Purity, expansion rate, receptor expression, and functions of eNK cells were determined over four weeks of culture. RESULTS: NK cell expansion was enhanced by short exposure of soluble IL-18 and IL-21 with K562-OX40L cells. Co-culture of NK cells with K562-OX40L-mbIL-18/-21 cells resulted in remarkable expansion of NK cells from HDs (9,860-fold) and MM patients (4,929-fold) over the 28-day culture period. Moreover, eNK cells showed increased expression of major activation markers and enhanced cytotoxicity towards target K562, U266, and RPMI8226 cells. CONCLUSIONS: Our data suggest that genetically engineered K562 cells expressing OX40L, mbIL-18, and mbIL-21 improve the expansion of NK cells, increase activation signals, and enhance their cytolytic activity towards MM cells.

Asthma, characterized by persistent inflammation and increased sensitivity of the airway, is the most common chronic condition among children. Novel, safe, and reliable treatment strategies are the focus of current research on pediatric asthma. Amygdalin, mainly present in bitter almonds, has anti-inflammatory and immunoregulatory potential, but its effect on asthma remains uninvestigated. Here, the impact of amygdalin on the thymic stromal lymphopoietin (TSLP)-- dendritic cell (DC)--OX40L axis was investigated. A BALB/c mouse model for allergic asthma was established using the ovalbumin-sensitization method. Amygdalin treatment was administered between days 21 and 27 of the protocol. Cell numbers and hematoxylin and eosin (H&E) staining in bronchoalveolar lavage fluid (BALF) were used to observe the impact of amygdalin on airway inflammation. TSLP, IL-4, IL-5, IL-13, and IFN-y concentrations were determined via Enzyme-linked immunosorbent assay (ELISA). TSLP, GATA-3, and T-bet proteins were measured using western blotting. Cell-surface receptor expression on DCs (MHC II, CD80, and CD86) was assessed via flow cytometry. OX40L mRNA and protein levels were detected using western blotting and qRT-PCR, respectively. Amygdalin treatment attenuated airway inflammation decreased BALF TSLP levels, inhibited DC maturation, restrained TSLP-induced DC surface marker expression (MHCII, CD80, and CD86), and further decreased OX40L levels in activated DCs. This occurred together with decreased Th2 cytokine levels (IL-4, IL-5, and IL-13) and GATA3 expression, whereas Th1 cytokine (IFN-y) levels and T-bet expression increased. Amygdalin thus regulates the Th1/Th2 balance through the TSLP--DC--OX40L axis to participate in inflammation development in the airways, providing a basis for potential allergic asthma treatments. [ABSTRACT FROM AUTHOR]

The epithelium-associated cytokine thymic stromal lymphopoietin (TSLP) can induce OX40L and CCL17 expression by myeloid dendritic cells (mDCs), which contributes to aberrant Th2-type immune responses. Herein, we report that such TSLP-induced Th2-type immune response can be effectively controlled by Dectin-1, a C-type lectin receptor expressed by mDCs. Dectin-1 stimulation induced STAT3 activation and decreased the transcriptional activity of p50-RelB, both of which resulted in reduced OX40L expression on TSLP-activated mDCs. Dectin-1 stimulation also suppressed TSLP-induced STAT6 activation, resulting in decreased expression of the Th2 chemoattractant CCL17. We further demonstrated that Dectin-1 activation was capable of suppressing ragweed allergen (Amb a 1)-specific Th2-type T cell response in allergy patients ex vivo and house dust mite allergen (Der p 1)-specific IgE response in non-human primates in vivo. Collectively, this study provides a molecular explanation of Dectin-1-mediated suppression of Th2-type inflammatory responses and suggests Dectin-1 as a target for controlling Th2-type inflammation.

We have previously shown GM-CSF derived bone-marrow dendritic cells (G-BMDCs) can induce the selective expansion of Tregs through the surface-bound molecule OX40L; however, the physiological role of this ex vivo derived DC subset remained to be elucidated. We determined GM-CSF administration to mice induced the generation of in vivo derived OX40L+ DCs, phenotypically similar to ex vivo OX40L+G-BMDCs, in the spleen, brachial lymph nodes and liver. The generation of OX40L+ DCs correlated with increased percentages of functionally suppressive Tregs in the spleen, brachial lymph nodes, and liver of GM-CSF treated mice. DCs from GM-CSF treated mice expanded Tregs in CD4+ T-cell co-cultures in an OX40L dependent manner, suggesting OX40L+ DCs may play a role in peripheral Treg homeostasis. Furthermore, comparing the transcriptome data of OX40L+ DCs to that of all immune cell types revealed OX40L+ DCs to be distinct from steady-state immune cells and, microarray analysis of OX40L+G-BMDCs and OX40L-G-BMDCs revealed higher expression of molecules that are associated with tolerogenic phenotype and could play important roles in the function of OX40L+ DCs. These findings suggest that OX40L+ DCs may represent a unique DC subset induced under inflammatory conditions that may play an essential role in maintaining Treg homeostasis.

Depending on age of acquisition, hepatitis B virus (HBV) can induce a cell-mediated immune response that results in either cure or progressive liver injury. In adult-acquired infection, HBV antigens are usually cleared, whereas in infancy-acquired infection, they persist. Individuals infected during infancy therefore represent the majority of patients chronically infected with HBV (CHB). A therapy that can promote viral antigen clearance in most CHB patients has not been developed and would represent a major health care advance and cost mitigator. Using an age-dependent mouse model of HBV clearance and persistence in conjunction with human blood and liver tissue, we studied mechanisms of viral clearance to identify new therapeutic targets. We demonstrate that age-dependent expression of the costimulatory molecule OX40 ligand (OX40L) by hepatic innate immune cells is pivotal in determining HBV immunity, and that treatment with OX40 agonists leads to improved HBV antigen clearance in young mice, as well as increased strength of T cell responses in young mice and adult mice that were exposed to HBV when they were young and developed a CHB serological profile. Similarly, in humans, we show that hepatic OX40L transcript expression is age-dependent and that increased OX40 expression on peripheral CD4+ T cells in adults is associated with HBV clearance. These findings provide new mechanistic understanding of the immune pathways and cells necessary for HBV immunity and identify potential therapeutic targets for resolving CHB.

Multiple research studies have demonstrated the efficacy of lactic acid bacteria in boosting both innate and adaptive immune responses. We have created a Lactococcus lactis variant that produces a modified combination protein with Fms-like tyrosine kinase 3 ligand and co-stimulator O × 40 ligand, known as HuFOLactis. The genetically modified variant was purposely created to activate T cells, NK cells, and DC cells in a laboratory setting. Furthermore, we explored the possibility of using the tumor-penetrating peptide iRGD to deliver HuFOLactis-activated immune cells to hard-to-reach tumor areas. Following brief stimulation with HuFOLactis, immune cell phenotypes and functions were assessed using flow cytometry. Confocal microscopy was employed to demonstrate the infiltrative and cytotoxic capabilities of iRGD-modified HuFOLactis-activated immune cells within tumor spheroids. The efficacy of iRGD modified HuFOLactis-activated immune cells against tumors was assessed in xenograft mouse models. HuFOLactis treatment resulted in notable immune cell activation, demonstrated by elevated levels of CD25, CD69, and CD137. Additionally, these activated immune cells showed heightened cytokine production and enhanced cytotoxicity against MKN45 cell lines. Incorporation of the iRGD modification facilitated the infiltration of HuFOLactis-activated immune cells into multicellular spheroids (MCSs). Additionally, immune cells activated by HuFOLactis and modified with iRGD, in combination with anti-PD-1 treatment, effectively halted tumor growth and prolonged survival in a mouse model of gastric cancer.

Background: Amlitelimab, a fully human nondepleting monoclonal antibody targeting OX40 ligand on antigen-presenting cells, could prevent T-cell-driven inflammation seen in atopic dermatitis (AD)., Objective: This trial evaluated the efficacy and safety of amlitelimab in adults with AD., Methods: In this 2-part, phase 2b, randomized, double-blinded placebo-controlled trial (NCT05131477), patients received subcutaneous amlitelimab every 4 weeks at doses of 250 mg with 500-mg loading dose, 250 mg, 125 mg, or 62.5 mg, or placebo for 24 weeks in Part 1 (1:1:1:1:1 randomization). In Part 2, clinical responders were reallocated 3:1 to withdraw amlitelimab or continue the previous dose regimen for 28 weeks. The primary end point was percent change in Eczema Area and Severity Index (EASI) from baseline to Week 16., Results: 390 and 190 patients enrolled in Part 1 and Part 2, respectively. Significant percent change decreases in EASI were observed with amlitelimab vs. placebo (P<.001). Clinical responses at Week 24 (Investigator Global Assessment 0/1 and/or EASI-75) were maintained at Week 52 in patients continuing or withdrawn from amlitelimab. In patients maintaining clinical response at Week 52 while off-treatment, >80% had serum amlitelimab concentrations below a 4-μg/mL threshold for several weeks prior to Week 52. Reductions in AD-related biomarkers during Part 1 were maintained through Part 2. Amlitelimab was well tolerated over 52 weeks., Conclusions: Amlitelimab treatment significantly reduced clinical and biomarker responses, and was well tolerated in adults with AD through Week 52. Sustained responses were observed in the majority of patients after amlitelimab withdrawal for 28 weeks., (Copyright © 2024. Published by Elsevier Inc.)

Xinxing Zhang,&ast; Zhichao Sun,&ast; Suyu Guo, Jiahui Zhang, Wenjing Gu, Zhengrong Chen, Li Huang Department of Pediatric Pulmonology, Children’s Hospital of Soochow University, Suzhou, Jiangsu, 215003, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Zhengrong Chen; Li Huang Tel/Fax +86-0512-80693588Email chen_zheng_rong@163.com; szdalv@163.comBackground: It was demonstrated that membrane-associated RING-CH 1 (March 1) might play an important role in the pathogenesis of asthma.Methods: The levels of mRNA and protein were measured by qRT-PCR and Western blot, respectively. Immunofluorescence assay was used to determine whether March1 co-locates with HDAC11. Co-immunoprecipitation was performed to examine the combination of proteins. Moreover, luciferase assay was used to measure the promoter activity of genes.Results: The mRNA and protein levels of both March1 and OX40 ligand (OX40L) were increased in the dendritic cells (DCs) from asthmatic children and asthmatic animals. Histone deacetylase 11 (HDAC11) protein was decreased in the DCs from asthmatic children and asthmatic model. Increasing of March1 or decreasing of March1 only affect the expression of HDAC11 in protein level. Besides, increasing of HDAC11 could inhibit OX40L expression, and decreasing of HDAC11 promoted OX40L expression in house dust mites (HDMs)-treated DCs. Increasing of HDAC11 notably reversed the promotion of March1 to OX40L expression. Our data further proved that March1 reduced the protein level of HDAC11 through inducing ubiquitination and degradation. HDAC11 combined with krüppel‐like factor 4 (KLF4) to decrease the activity of OX40L gene promoter, thus to downregulate the level of OX40L.Conclusion: Overall, our data showed that HDAC11 promoted KLF4-dependent OX40L decreasing. However, March1 promoted OX40L expression through enhancing the ubiquitination and degradation of HDAC11 and subsequent blocking the inhibition of HDAC11 to OX40L.Keywords: asthma, march1 regulating OX40 ligand, ubiquitination, histone deacetylase 11, OX40 ligand

Body's defence mechanism has ability to combat tumour cells but tumour cells can circumvent immune system in order to flourish. Therefore, current research focuses on reinvigorating immune system to combat against extensive range of human malignancies through immunotherapy. Recently, immuno-therapy has demonstrated beneficial outcomes in cancers treatment but the main drawbacks are primary and acquired resistance to the therapeutic agents and immune-related toxicities. Therefore, novel immune therapies are direly required. Co-stimulatory molecules such as TNF Receptor Superfamily Member 4 (OX40, CD134) and its ligand TNF Superfamily Member 4 (CD252, OX40L) are expressed on different immune cells. The mutual interaction between OX40 and its ligand (OX40/OX40L) decreases the functional capacity of immunosuppression offered by regulatory T cells (Tregs) and induces the proliferation of T cells against specific antigen enhancing the immune response. Many clinical trials are focusing on OX40/OX40L therapeutic agents to find out whether they have therapeutic effect on cancer treatment. The initial phase trials result of OX40 and its ligands focusing therapeutic agents are encouraging but still not sufficient. This review will concentrate on the cellular and molecular pathways of OX40-mediated T-cell co-stimulation, the expression of OX40 and OX40L in tumours, the implications of their interactions and their under-or over-expression patterns, with particular focus on the function of OX40 in tumours of different origins. Finally, we discuss results of clinical trials of OX40 and OX40L directed pharmacotherapy and the lacunae that need to be filled.

We have previously shown that OX40L/OX40 interaction is critical for TCR-independent selective proliferation of Foxp3+ Tregs, but not Foxp3- effector T-cells (Teff), when CD4+ T-cells are co-cultured with GM-CSF derived bone marrow dendritic cells (G-BMDCs). Events downstream of OX40L/OX40 interaction in Tregs responsible for this novel mechanism are not understood. Earlier, OX40L/OX40 interaction has been shown to stimulate CD4+ T-cells through the formation of a signalosome involving TRAF2/PKC-Ѳ leading to NF-kB activation. In this study, using CD4+ T-cells from WT and OX40-/- mice we first established that OX40 mediated activation of NF-kB was critical for this Treg proliferation. Although CD4+ T-cells from PKC-Ѳ-/- mice were also defective in G-BMDC induced Treg proliferation ex vivo, this defect could be readily corrected by adding exogenous IL-2 to the co-cultures. Furthermore, by treating WT, OX40-/-, and PKC-Ѳ-/- mice with soluble OX40L we established that OX40L/OX40 interaction was required and sufficient to induce Treg proliferation in vivo independent of PKC-Ѳ status. Although PKC-Ѳ is dispensable for TCR-independent Treg proliferation per se, it is essential for optimum IL-2 production by Teff cells. Finally, our findings suggest that OX40L binding to OX40 likely results in recruitment of TRAF1 for downstream signalling.

BackgroundRhinovirus infection at an early age has been associated with development of asthma, but how rhinovirus influences the immune response is not clear.ObjectiveTolerance to inhaled antigen is mediated through induction of regulatory T (Treg) cells, and we examined whether rhinovirus infection of the respiratory tract can block airway tolerance by modulating Treg cells.MethodsThe immune response to intranasal ovalbumin in mice was assessed with concomitant infection with RV1B, and the factors induced in vivo were compared with those made by human lung epithelial cells infected in vitro with RV16.ResultsRV1B infection of mice abrogated tolerance induced by inhalation of soluble ovalbumin, suppressing the normal generation of forkhead box protein 3-positive Treg cells while promoting TH2 cells. Furthermore, RV1B infection led to susceptibility to asthmatic lung disease when mice subsequently re-encountered aeroantigen. RV1B promoted early in vivo expression of the TNF family protein OX40 ligand on lung dendritic cells that was dependent on the innate cytokine thymic stromal lymphopoietin (TSLP) and also induced another innate cytokine, IL-33. Inhibiting each of these pathways allowed the natural development of Treg cells while minimizing TH2 differentiation and restored tolerance in the face of RV1B infection. In accordance, RV16 infection of human lung epithelial cells upregulated TSLP and IL-33 expression.ConclusionsThese results suggest that infection of the respiratory epithelium with rhinovirus can antagonize tolerance to inhaled antigen through combined induction of TSLP, IL-33, and OX40 ligand and that this can lead to susceptibility to asthmatic lung inflammation.

Monoclonal antibodies targeting immune checkpoints have been widely applied in gastrointestinal cancer immunotherapy. However, systemic administration of various monoclonal antibodies does not often result in sustained effects in reversing the immunosuppressive tumor microenvironment (TME), which may be due to the spatiotemporal dynamic changes of immune checkpoints. Herein, we reported a novel immune checkpoint reprogramming strategy for gastrointestinal cancer immunotherapy. It was achieved by the sequential delivery of siPD-L1 (siRNA for programmed cell death ligand 1) and pOX40L (plasmid for OX40 ligand), which were complexed with two cationic polymer brush-grafted carbon nanotubes (dense short (DS) and dense long (DL)) designed based on the structural characteristics of nucleic acids and brush architectures. Upon administrating DL/pOX40L for the first three dosages, then followed by DS/siPD-L1 for the next three dosages to the TME, it upregulated the stimulatory checkpoint OX40L on dendritic cells (DCs) and downregulated inhibitory checkpoint PD-L1 on tumor cells and DCs in a sequential reprogramming manner. Compared with other combination treatments, this sequential strategy drastically boosted the DCs maturation, and CD8 + cytotoxic T lymphocytes infiltration in tumor site. Furthermore, it could augment the local antitumor response and improve the T cell infiltration in tumor-draining lymph nodes to reverse the peripheral immunosuppression. Our study demonstrated that sequential nucleic acid delivery strategy via personalized nanoplatforms effectively reversed the immunosuppression status in both tumor microenvironment and peripheral immune landscape, which significantly enhanced the systemic antitumor immune responses and established an optimal immunotherapy strategy against gastrointestinal cancer., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Background: OX40 and OX40 ligand (OX40L), as essential immune checkpoint (IC) modulators, are highly correlated with cancer immunity regulation as well as tumor microenvironment (TME). Immunotherapy showed outstanding advantages in small-cell lung cancer (SCLC) therapy. However, functions and clinical significance of OX40 and OX40L in SCLC were not clear yet. Materials and Methods: SCLC samples of 143 patients were collected for immunohistochemistry (IHC) or whole-exome sequencing (WES). We comprehensively explored the expression and mutation of OX40/OX40L in SCLC, and systematically linked OX40/OX40L with TME. Results: The expression of OX40/OX40L on tumor cells and tumor-infiltrating lymphocytes (TILs) was found in the IHC cohort and verified in other cohorts with SCLC tissues and cell lines. The results showed co-expression patterns among OX40/OX40L, other ICs, and T-cell markers. The WES data suggested that OX40/OX40L mutation is rare in SCLC (<5%). Patients with positive OX40 protein expression on TILs showed substantially higher recurrence-free survival than those with negative expression (p=0.009). The external dataset also indicated that high OX40/OX40L expression was correlated with better prognosis [overall survival: OX40, p<0.001; OX40L, p=0.019]. Importantly, activation of immunity and high infiltration of CD4(+) and CD8(+) T cells were observed in the high OX40/OX40L expression group. Conclusions: Collectively, this work highlighted the significance of OX40 and OX40L in prognosis and TME cell infiltration characterization of SCLC. Evaluating the OX40/OX40L-expression levels of individual patients with SCLC might contribute to guiding more precise therapy. [ABSTRACT FROM AUTHOR]

Objective To construct OX40Ig genetically modified rat adipose derived mesenchymal stromal cells (ADSCsOX40Ig), and to investigate its immunomodulatory effect on lymphocytes in vitro. Methods The fusion gene expression vector of pcDNA3.1(+)/OX40Ig was constructed and transfected into rat ADSCs by nuclear transfection. The expression of OX40Ig in ADSCs was measured by Western blot assay. The transfected and untransfected ADSCs were co-cultured with rat allogeneic lymphocytes, and were divided into the control group (responder + stimulator), the ADSCs (responder+stimulator+ ADSCs) group, and the ADSCsOX40Ig (responder+stimulator+ADSCsOX40Ig) group. The proliferatory activity of lymphocytes was detected by MTT method. Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect interferon (IFN)-γ, interleukin (IL) -10 and transforming growth factor (TGF) - β mRNA levels in lymphocytes. Results The sequencing confirmed that pcDNA3.1(+)/OX40Ig plasmid was successfully constructed, and OX40Ig was highly expressed in transfected ADSCs. Compared with ADSCs, the proliferation inhibition rate of allogeneic lymphocytes was significantly increased in ADSCsOX40Ig (P＜0.05). The expression levels of IFN-γ mRNA decreased gradually, but the expression levels of IL-10 and TGF-β mRNA increased gradually in the control, ADSCs and ADSCsOX40Ig groups (P＜0.05). Conclusion Compared with ADSCs along, ADSCsOX40Ig has better immunomodulatory effects on allogeneic lymphocytes. [ABSTRACT FROM AUTHOR]

BackgroundOX40 and OX40 ligand (OX40L), as essential immune checkpoint (IC) modulators, are highly correlated with cancer immunity regulation as well as tumor microenvironment (TME). Immunotherapy showed outstanding advantages in small-cell lung cancer (SCLC) therapy. However, functions and clinical significance of OX40 and OX40L in SCLC were not clear yet.Materials and MethodsSCLC samples of 143 patients were collected for immunohistochemistry (IHC) or whole-exome sequencing (WES). We comprehensively explored the expression and mutation of OX40/OX40L in SCLC, and systematically linked OX40/OX40L with TME.ResultsThe expression of OX40/OX40L on tumor cells and tumor-infiltrating lymphocytes (TILs) was found in the IHC cohort and verified in other cohorts with SCLC tissues and cell lines. The results showed co-expression patterns among OX40/OX40L, other ICs, and T-cell markers. The WES data suggested that OX40/OX40L mutation is rare in SCLC (

Objective To investigate the effects of shikonin on thymic stromal lymphopoietin (TSLP)/OX40 ligand (OX40L) pathway and balance of helper T cell 1 (Th1)/helper T cell 2 (Th2) in mice with atopic dermatitis (AD). Methods AD mouse model was established by topical application of 2,4-dinitrofluorobenzene (DNFB).Model mice were randomly divided into model group, low shikonin group (10 mg/kg), medium shikonin group (20 mg/kg), high shikonin group (30 mg/kg), and the positive control group (prednisolone,10 mg/kg) by random number table method, with 15 mice in each group. Another 15 mice were used as the normal control group. The drug groups were given the corresponding drugs by gavage, with the volume of 10 mL/kg. The model group and the normal control group were given the same volume of solvent by gavage, once a day for 15 days. The scratch behavior of mice was observed on the 5th, 10th and 15th day after administration. The condition of skin lesions was evaluated on the 7th and 15th days after administration. At the end of the last administration, hematoxylin eosin (HE) staining was used to observe the histopathological changes of skin lesions. The ratio of Th1/Th2 cells in peripheral blood of mice was detected by flow cytometry. Levels of TSLP, tumor necrosis factor (TNF)-α, interleukin (IL)-4 and γ-interferon (IFN-γ) were detected by enzyme linked immunosorbent assay (ELISA). Western blot assay was used to detect the protein expression levels of TSLP and OX40L. Results Compared with the normal control group, the scratching behavior increased and the skin inflammatory injury aggravated in model group. There were dense infiltration of inflammatory cells in the epidermis and dermis in the model group, and the epidermis was thickened. The blood levels of Th2 cells, TSLP, TNF-α and IL-4, and the protein expression levels of TSLP and OX40L in skin lesions were significantly higher (P＜0.05), and the levels of Th1 cells, IFN-γ and Th1/Th2 ratio were significantly lower (P＜0.05). Compared with the model group, the scratching behavior were decreased on the 5th, the 10th and the 15th day, and the skin inflammatory injury decreased on the 7th and the 15th day in turn in low, middle and high dose shikonin groups (P＜0.05). After the last administration, the inflammatory cell infiltration and epidermal thickening in the epidermis and dermis of mice decreased in turn in low, medium and high dose shikonin groups. The scratching behavior, the degree of skin inflammatory injury, the levels of Th2 cells, TSLP, TNF-α and IL-4, and the protein expression levels of TSLP and OX40L decreased in turn (P＜ 0.05), and the levels of Th1 cells, IFN-γ and Th1/Th2 ratio increased in turn (P＜0.05). Conclusion Shikonin may maintain the balance of Th1/Th2 cells in peripheral blood and improve skin inflammatory injury of AD mice by inhibiting TSLP/OX40L pathway. [ABSTRACT FROM AUTHOR]

Body‟s defence mechanism has ability to combat tumour cells but tumour cells can circumvent immune system in order to flourish. Therefore, current research focuses on reinvigorating immune system to combat against extensive range of human malignancies through immunotherapy. Recently, immuno-therapy has demonstrated beneficial outcomes in cancers treatment but the main drawbacks are primary and acquired resistance to the therapeutic agents and immune-related toxicities. Therefore, novel immune therapies are direly required. Co-stimulatory molecules such as TNF Receptor Superfamily Member 4 (OX40, CD134) and its ligand TNF Superfamily Member 4 (CD252, OX40L) are expressed on different immune cells. The mutual interaction between OX40 and its ligand (OX40/OX40L) decreases the functional capacity of immunosuppression offered by regulatory T cells (Tregs) and induces the proliferation of T cells against specific antigen enhancing the immune response. Many clinical trials are focusing on OX40/OX40L therapeutic agents to find out whether they have therapeutic effect on cancer treatment. The initial phase trials result of OX40 and its ligands focusing therapeutic agents are encouraging but still not sufficient. This review will concentrate on the cellular and molecular pathways of OX40-mediated T-cell co-stimulation, the expression of OX40 and OX40L in tumours, the implications of their interactions and their under-or over-expression patterns, with particular focus on the function of OX40 in tumours of different origins. Finally, we discuss results of clinical trials of OX40 and OX40L directed pharmacotherapy and the lacunae that need to be filled. [ABSTRACT FROM AUTHOR]

Adoptive natural killer (NK) cell transfer has been demonstrated to be a promising immunotherapy approach against malignancies, but requires the administration of sufficient activated cells for treatment effectiveness. However, the paucity of clinical-grade to support the for large-scale cell expansion limits its feasibility. Here we developed a feeder-based NK cell expansion approach that utilizes OX40L armed NK-92 cell with secreting neoleukin-2/15 (Neo-2/15), a hyper-stable mimetic with a high affinity to IL-2Rβγ. The novel feeder cells (NK92-Neo2/15-OX40L) induced the expansion of NK cells with a 2180-fold expansion (median; 5 donors; range, 1767 to 2719) after 21 days of co-culture without added cytokines. These cells were highly cytotoxic against Raji cells and against several solid tumors in vivo. Mechanistically, NK92-Neo2/15-OX40L induced OX40 and OX40L expression on expanded NK cells and promoted the OX40-OX40L positive feedback loop, thus boosting NK cell function. Our data provided a novel NK cell expansion mechanism and insights into OX40-OX40L axis regulation of NK cell expansion.

Introduction: Osteoarthritis (OA) is a common musculoskeletal disease characterized by pain, stiffness, limited activity, occasional effusion, and local inflammation. MiR-146 is one of the noncoding RNA closely related to OA, but the role of miR-146 in OA remains controversial. The tumour necrosis factor receptor OX40 is activated by its cognate ligand OX40L (TNFSF4) and functions as a T-cell costimulatory molecule. The T-cell functions, including cytokine production, expansion, and survival, are enhanced by the OX40 costimulatory signals. Methods: We established an inflammatory model of condylar chondrocytes induced by IL-1β and TNF-α and detected the expression of miRNA by miRNA sequencing. Then, cell transfection was used to study the role of miR146a-5p in OA. The Kyoto Encyclopedia of Genes and Genomes (KEGG) and database analysis were used to screen out potential target genes of miR-146a-5p. A dual luciferase activity assay tested whether ox40l is the target gene of miR-146a-5p. Results: MiR-146a-5p and OX40L was upregulated after induced by IL-1β and TNF-α, miR-146a-5p reduced the production of inflammatory factors but had no effect on chondrophenotypic factors, and ox40l was targeted by miR-146a-5p. Conclusion: OX40L and miR-146a-5p of condylar chondrocytes in the inflammatory environment (induced by IL-1β and TNF-α) were significantly increased, miR-146a-5p is a protective factor in the inflammatory response, which can reduce the production of inflammatory factors, and miR-146a-5p may regulate T-cell-mediated immunity through targeting of ox40l in OA. [ABSTRACT FROM AUTHOR]

Adoptive natural killer (NK) cell transfer has been demonstrated to be a promising immunotherapy approach against malignancies, but requires the administration of sufficient activated cells for treatment effectiveness. However, the paucity of clinical-grade to support the for large-scale cell expansion limits its feasibility. Here we developed a feeder-based NK cell expansion approach that utilizes OX40L armed NK-92 cell with secreting neoleukin-2/15 (Neo-2/15), a hyper-stable mimetic with a high affinity to IL-2Rβγ. The novel feeder cells (NK92-Neo2/15-OX40L) induced the expansion of NK cells with a 2180-fold expansion (median; 5 donors; range, 1767 to 2719) after 21 days of co-culture without added cytokines. These cells were highly cytotoxic against Raji cells and against several solid tumors in vivo. Mechanistically, NK92-Neo2/15-OX40L induced OX40 and OX40L expression on expanded NK cells and promoted the OX40-OX40L positive feedback loop, thus boosting NK cell function. Our data provided a novel NK cell expansion mechanism and insights into OX40-OX40L axis regulation of NK cell expansion. [ABSTRACT FROM AUTHOR]

This study sought to compare the behavior of Treg subsets displaying different coexpression patterns of Neuropilin-1 (Nrp1) and Helios, under the influence of gut stress unrelated to hematopoietic stem cell transplantation, pretransplantation conditioning, and posttransplant gastrointestinal acute graft versus host disease (GI-aGvHD). Host CD4 + /CD25 hi /Foxp3 + Treg cells, identified by flow cytometry, were isolated from various tissues of mice affected by these stressors. Expression of CD25, CTLA-4, CD39, OX40, integrin-β7, LAG3, TGFβ/LAP, granzyme-A, -B, and interleukin-10 was compared in four Treg subsets displaying Helios or Nrp1 only, both or none. Fluorescence-activated cell sorter-sorted Treg subsets, displaying markers affected in a conditioning- and GI-aGVHD-restricted manner, were further investigated by transcriptome profiling and T-cell suppression assays. We found that conditioning by irradiation greatly diminished the relative frequency of Helios + /Nrp1 + Treg, shifting the balance toward Helios - /Nrp1 - Treg in the host. Upregulation of integrin-β7 and OX40 occurred in GI-aGvHD-dependent manner in Helios + /Nrp1 + cells but not in Helios - /Nrp1 - Treg. Sorted Treg subsets, confirmed to overexpress Nrp1, Helios, OX40, or integrin-β7, displayed superior immunosuppressive activity and enrichment in activation-related messenger RNA transcripts. Our data suggest that conditioning-induced shrinkage of the Nrp1 + /Helios + Treg subset may contribute to the development of GI-GvHD by impairing gut homing and decreasing the efficiency of Treg-mediated immunosuppression., (© 2024 The Authors. European Journal of Immunology published by Wiley‐VCH GmbH.)

Background: The efficacy of neoadjuvant chemo-immunotherapy (NAT) in esophageal squamous cell carcinoma (ESCC) is challenged by the intricate interplay within the tumor microenvironment (TME). Unveiling the immune landscape of ESCC in the context of NAT could shed light on heterogeneity and optimize therapeutic strategies for patients., Methods: We analyzed single cells from 22 baseline and 24 post-NAT treatment samples of stage II/III ESCC patients to explore the association between the immune landscape and pathological response to neoadjuvant anti-PD-1 combination therapy, including pathological complete response (pCR), major pathological response (MPR), and incomplete pathological response (IPR)., Results: Single-cell profiling identified 14 major cell subsets of cancer, immune, and stromal cells. Trajectory analysis unveiled an interesting link between cancer cell differentiation and pathological response to NAT. ESCC tumors enriched with less differentiated cancer cells exhibited a potentially favorable pathological response to NAT, while tumors enriched with clusters of more differentiated cancer cells may resist treatment. Deconvolution of transcriptomes in pre-treatment tumors identified gene signatures in response to NAT contributed by specific immune cell populations. Upregulated genes associated with better pathological responses in CD8 + effector T cells primarily involved interferon-gamma (IFNγ) signaling, neutrophil degranulation, and negative regulation of the T cell apoptotic process, whereas downregulated genes were dominated by those in the immune response-activating cell surface receptor signaling pathway. Natural killer cells in pre-treatment tumors from pCR patients showed a similar upregulation of gene expression in response to IFNγ but a downregulation of genes in the neutrophil-mediated immunity pathways. A decreased cellular contexture of regulatory T cells in ESCC TME indicated a potentially favorable pathological response to NAT. Cell-cell communication analysis revealed extensive interactions between CCL5 and its receptor CCR5 in various immune cells of baseline pCR tumors. Immune checkpoint interaction pairs, including CTLA4-CD86, TIGIT-PVR, LGALS9-HAVCR2, and TNFSF4-TNFRSF4, might serve as additional therapeutic targets for ICI therapy in ESCC., Conclusions: This pioneering study unveiled an intriguing association between cancer cell differentiation and pathological response in esophageal cancer patients, revealing distinct subgroups of tumors for which neoadjuvant chemo-immunotherapy might be effective. We also delineated the immune landscape of ESCC tumors in the context of clinical response to NAT, which provides clinical insights for better understanding how patients respond to the treatment and further identifying novel therapeutic targets for ESCC patients in the future., (© 2024. The Author(s).)

In spite of the well-known clustering of multiple autoimmune disorders in families, analyses of specific shared genes and polymorphisms between systemic lupus erythematosus (SLE) and other autoimmune diseases (ADs) have been limited. Therefore, we comprehensively tested autoimmune variants for association with SLE, aiming to identify pleiotropic genetic associations between these diseases. We compiled a list of 446 non-Major Histocompatibility Complex (MHC) variants identified in genome-wide association studies (GWAS) of populations of European ancestry across 17 ADs. We then tested these variants in our combined Caucasian SLE cohorts of 1,500 cases and 5,706 controls. We tested a subset of these polymorphisms in an independent Caucasian replication cohort of 2,085 SLE cases and 2,854 controls, allowing the computation of a meta-analysis between all cohorts. We have uncovered novel shared SLE loci that passed multiple comparisons adjustment, including the VTCN1 (rs12046117, P = 2.02×10(-06)) region. We observed that the loci shared among the most ADs include IL23R, OLIG3/TNFAIP3, and IL2RA. Given the lack of a universal autoimmune risk locus outside of the MHC and variable specificities for different diseases, our data suggests partial pleiotropy among ADs. Hierarchical clustering of ADs suggested that the most genetically related ADs appear to be type 1 diabetes with rheumatoid arthritis and Crohn's disease with ulcerative colitis. These findings support a relatively distinct genetic susceptibility for SLE. For many of the shared GWAS autoimmune loci, we found no evidence for association with SLE, including IL23R. Also, several established SLE loci are apparently not associated with other ADs, including the ITGAM-ITGAX and TNFSF4 regions. This study represents the most comprehensive evaluation of shared autoimmune loci to date, supports a relatively distinct non-MHC genetic susceptibility for SLE, provides further evidence for previously and newly identified shared genes in SLE, and highlights the value of studies of potentially pleiotropic genes in autoimmune diseases.

Objective To investigate the expression and clinical significance of OX40/ OX40L (CD134/ CD134L) in CD4+T cells, CDS+ T cells, monocytes, and B lymphocytes in peripheral blood of patients with autoimmune hepatitis (AIH), primary biliary cholangitis (PBC), and their overlap syndrome before and after standardized treatment. Methods A total of 74 patients with AIH, PBC, and their overlap syndrome who were diagnosed in Subei People' s Hospital of Jiangsu from August 2015 to August 2019 were enrolled, and according to related diagnostic criteria, they were divided into AIH group (group A) with 29 patients, PBC group (group P) with 26 patients, and overlap syndrome group ( group C) with 19 patients. A healthy control group with 30 individuals was also established. Peripheral blood samples were collected before and after standardized treatment to measure the expression of OX40/OX40L on the surface of peripheral blood cells by immunofluorescence flow cytometry, and the expression of OX40/OX40L was compared before and after treatment and between the three groups and the healthy control group to investigate its clinical significance. A one - way analysis of variance was used for comparison between multiple groups, and the least significant difference t - test was used for further comparison between two groups; the paired t - test was used for comparison of paired samples between two groups. Results There were no significant differences in sex composition and age composition between the three groups ( P > 0.05). Before treatment, the positive rate of OX40 in peripheral blood CD4+ T cells gradually increased in groups A, P, and C, and groups A, P, and C had a significantly higher positive rate of OX40 than the control group (14.80% ± 4.99% / 17.11% ± 2.71 % /25.18% ± 5.55% vs 6.67% ± 2.26%, F = 14. 823, P <0.001); groups A, P, and C had a significantly higher positive rate of OX40 in CD8+ T cells than the control group (4.86% ± 1.54% / 6.40% ± 1.88% /7.33% ± 2.12% vs 4.09% ± 2.69%, F = 5.486, P <0.001); the positive rate of OX40L in CD14+ monocytes was 19.84% ± 6.11% in group A, 21.17% ± 4.35% in group P, 29.13% ± 6.32% in group C, and 4.86% ±2.34% in the control group, and there was a significant difference between groups (F = 17.004, P <0.001); the positive rate of OX40L in CD19+ B cells was 17.62% ± 3.86% in group A, 14.75% ± 4.32% in group P, 10.13% ± 2.56% in group C, and 4. 50% ± 1.38% in the control group, showing a trend of gradual reduction, and groups A, P, and C had a significantly higher positive rate than the control group (F = 12.221, P < 0.001). After treatment, the positive rate of OX40 in CD8+ T cells decreased significantly to a similar level as the control group, and there was no significant difference between groups (F = 0. 731, P = 0.538). For the other three types of cells, although there were varying degrees of reduction in the positive rate of OX40/ OX40L after treatment, groups A, P, and C still had a significantly higher positive rate than the control group; in CD4+ T cells, the positive rate of OX40 was 11.00% ± 1.98% in group A, 13.72% ± 1.03% ingroup P, 19.72% ±3.47% ingroupC, and6.67% ±2.26% in the control group, and groups A, P, and C had a significantly higher positive rate than the control group (F = 11.365, P < 0. 001); in CD14+ monocytes, the positive rate of OX40L was 11.82% ± 2.23% in group A, 15.19% ± 4.42% in group P, 24.51% ± 4.09% in group C, and 4.86% ± 2.34% in the control group, and groups A, P, and C had a significantly higher positive rate than the control group (F = 13.748, P<0.001); in CD19+ B cells, the positive rate of OX40L was 9.09% ±3.25% in group A, 6.81% ±2. 20% in group P, 7.48% ± 2.85% in group C, and 4.50% ± 1.38% in the control group, and groups A, P, and C had a significantly higher positive rate than the control group (F = 8.052, P < 0.001). Groups A, P, and C had significant reductions in the expression of OX40/ OX40L in peripheral blood CD4+ T cells, CD8+ T cells, CD14+ monocytes, and CD19+ B lymphocytes after treatment (all P < 0.05). Conclusion The expression of OX40/ OX40L in peripheral blood increases in patients with AIH, PBC, and their overlap syndrome and decreases after treatment, indicating that the OX40/ OX40L pathway is involved in the pathogenesis of the above diseases, and the role of OX40 on the surface of CD8+ T cells may better reflect the treatment outcome. [ABSTRACT FROM AUTHOR]

Introduction: Interaction of OX40 and OX40 ligand (OX40L) is associated with immune activation. OX40-OX40L axis is also suggested to play a role in immunity against several solid malignancies. Objective: In this study, serum OX40 and OX40L levels in patients with advanced lung adenocarcinoma were assessed and their correlation with survival and clinicopathologic parameters was determined. Methods: Serum samples were collected from patients with advanced lung adenocarcinoma, then OX40 and OX40L were quantified via enzyme-linked immunosorbent assay. Immunohistochemical (IHC) analysis of OX40 and OX40L in resected primary lesions was also performed. The association between OX40 and OX40L levels and clinicopathologic status and patient survival was retrospectively analyzed. Results: A total of 56 patients were analyzed. Median serum OX40 and OX40L levels were 156.2 pg/mL and 186.6 pg/mL, respectively. IHC analysis in 5 patients indicated high positivity of OX40 in tumor-infiltrating lymphocytes and of OX40L in tumor cells in mucinous adenocarcinoma. Patients with a high OX40 level (≥152.2 pg/mL) had poorer prognosis than those with a low serum OX40 level (median survival, 7.36 vs. 21.19 months, respectively, p = 0.04). Patients with a high OX40L level (≥207.3 pg/mL) had poorer prognosis than those with a low serum OX40L level (median survival, 7.36 vs. 14.26 months, respectively, p = 0.04). In the subset of patients treated with immune checkpoint inhibitors (ICIs) (n = 12), those with a high OX40L level were found to have longer survival from ICI initiation than those with a low OX40L level (p = 0.023). Conclusions: High OX40 and OX40L levels are associated with poor prognosis and may reflect the immune-exhausted status against lung adenocarcinoma. [ABSTRACT FROM AUTHOR]

Acute or lymphomatous type adult T cell leukemia/lymphoma (ATLL) is an aggressive hematopoietic malignancy with poor prognosis. We previously reported that programmed cell death ligand 1 (PD-L1) expression could predict ATLL outcomes. However, the roles of other immune checkpoint molecules remain largely unknown in ATLL. Our aim in this study was to explore the clinicopathological impacts of immune checkpoint molecules in ATLL. Immunohistochemistry was performed in 69 ATLL patients with antibodies against the following: PD-L1, programmed cell death ligand 2 (PD-L2), OX40, OX40 ligand (OX40L), CD137, CD137 ligand (CD137L), Galectin-9, T cell immunoglobulin mucin-3 (Tim-3), cytotoxic T lymphocyte associated protein-4 (CTLA-4), lymphocyte activating-3 (LAG-3), CD80, CD86, glucocorticoid-induced tumor necrosis factor receptor-related protein (GITR), GITR ligand (GITRL), and programmed death-1 (PD-1). Immune checkpoint molecules were variably expressed on neoplastic and/or microenvironmental cells. Expression of PD-1, OX40L, Galectin-9, and PD-L1 was nearly mutually exclusive on neoplastic cells, suggesting that immune checkpoint pathways differ in patients. Microenvironmental expression of PD-L1, OX40L, and Tim-3 was significantly associated with better overall survival (log-rank test; P =0.0004, 0.0394, and 0.0279, respectively). Univariate and multivariate analyses with clinical prognostic factors identified microenvironmental expression of PD-L1 and OX40L, and age (> 70 years) as significant prognostic factors. This is the first comprehensive analysis of ATLL immune checkpoint molecules. Our results may provide information on new therapeutic strategies in ATLL. [ABSTRACT FROM AUTHOR]

Background: Natural Killer (NK) cell-based immunotherapy used to treat cancer requires the adoptive transfer of a large number of activated NK cells. Here, we report a new effective method to expand human NK cells ex vivo using K562 cells genetically engineered (GE) to express OX40 ligand (K562-OX40L) in combination with a short exposure to soluble IL-21. In addition, we describe a possible mechanism of the NK cell expansion through the OX40 receptor-OX40 ligand axis which is dependent on NK cell homotypic interaction.Methods: K562-OX40L cells were generated by lentiviral transduction and were used as feeder cells to expand and activate NK cells from PBMCs in the presence of IL-2/IL-15. Soluble IL-21 was also added in various concentrations only once at the beginning of the culture. NK cells were expanded for 4–5 weeks, and the purity, expansion rate, phenotype and function (cytotoxicity, antibody-dependent cell-mediated cytotoxicity (ADCC), cytokine production, CD107a degranulation) of these expanded NK cells were compared to those generated by using K562 feeder cells.Results: The culture of NK cells with K562-OX40L cells in combination with the transient exposure to IL-21 highly enhanced NK cell expansion to approximately 2,000-fold after 4 weeks of culture, compared to a 303-fold expansion using the conventional K562 cells. Mechanistically, the OX40-OX40L axis between the feeder cells and NK cells as well as the homotypic interaction between NK cells through the OX40-OX40L axis were both necessary for NK cell expansion. The short exposure of NK cells to IL-21 had a synergistic effect with OX40 signaling for NK cell expansion. Apart from their enhanced expansion, NK cells grown with K562-OX40L feeder cells were similar to those grown with conventional K562 cells in regard to the surface expression of various receptors, cytotoxicity, ADCC, cytokine secretion, and CD107 degranulation.Conclusion: Our data suggest that OX40 ligand is a potent co-stimulant for the robust expansion of human NK cells and the homotypic NK cell interactions through the OX40-OX40L axis is a mechanism of NK cell expansion.

Sclerotic-type cutaneous chronic graft-versus-host disease is a severe complication of allogeneic hematopoietic stem cell transplantation, with profound morbidity. A dearth of effective, targeted treatment options necessitates further investigation into the molecular mechanisms underlying this T-cell-mediated disease. In this study, we compared the transcriptome in skin biopsies from pediatric and young adult (aged <25 years) patients with sclerotic-type cutaneous chronic graft-versus-host disease (n = 7) with that in demographically matched healthy controls (n = 8) and patients with atopic dermatitis (n = 10) using RNA sequencing with RT-PCR and immunohistochemistry validation. Differential expression was defined as fold change > 1.5 and false discovery rate < 0.05. Sclerotic-type cutaneous chronic graft-versus-host disease exhibited strong and significant T helper (Th)1 skewing through key related cytokines and chemokines (CXCL9/10/11, IFNG/IFN-γ, STAT1/signal transducer and activator of transcription 1). Several markers related to the TSLP-OX40 axis were significantly upregulated relative to those in both controls and lesional atopic dermatitis, including TNFSF4/OX40L, TSLP, and IL33, as well as fibroinflammatory signatures characterized in a prior study in systemic sclerosis. Gene set variation analysis reflected marker-level findings, showing the greatest enrichment of the Th1 and fibroinflammatory pathways, with no global activation identified in Th2 or Th17/Th22. Cell-type deconvolution revealed a significant representation of macrophages and vascular endothelial cells. Sclerotic-type cutaneous chronic graft-versus-host disease in young patients may therefore be characterized by strong Th1-related upregulation with a unique TSLP-OX40 signature, suggesting new therapeutic avenues for this devastating disease., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Disulfidptosis is a condition where dysregulated NAPDH levels and abnormal accumulation of cystine and other disulfides occur in cells with high SLC7A11 expression under glucose deficiency. This disrupts normal formation of disulfide bonds among cytoskeletal proteins, leading to histone skeleton collapse and triggering cellular apoptosis. However, the correlation between disulfidptosis and immune responses in relation to glioblastoma survival rates and immunotherapy sensitivity remains understudied. Therefore, we utilized The Cancer Genome Atlas and The Chinese Glioma Genome Atlas to identify disulfidptosis-related immune checkpoint genes and established an overall survival (OS) prediction model comprising six genes: CD276, TNFRSF 14, TNFSF14, TNFSF4, CD40, and TNFRSF18, which could also be used for predicting immunotherapy sensitivity. We identified a cohort of glioblastoma patients classified as high-risk, which exhibited an upregulation of angiogenesis, extracellular matrix remodeling, and epithelial-mesenchymal transition as well as an immunosuppressive tumor microenvironment (TME) enriched with tumor associated macrophages, tumor associated neutrophils, CD8  +  T-cell exhaustion. Immunohistochemical staining of CD276 in 144 cases further validated its negative correlation with OS in glioma. Disulfidptosis has the potential to induce chronic inflammation and an immunosuppressive TME in glioblastoma., (© 2024. The Author(s).)

The identification of new approaches and intervention targets for the treatment of AR is urgently needed. We aimed to investigate the effect of blocking the OX40/OX40L signaling pathway by small interfering RNA (siRNA) on ovalbumin (OVA)-induced AR in a mouse model. After establishment of the AR model, the mice were interfered by siRNA-OX40L (experimental group), siRNA-C (negative control group), or PBS (control group). Nose scratching, sneezing and nasal discharge were observed. OX40L mRNA and protein and the IL-5, TNF-α, regulatory T cell (Treg) -specific marker Foxp3, and eosinophil (EOS) levels were analyzed. The numbers of nose scratching and sneezing were significantly lower in the siRNA-OX40L-treated group (p <0.05). After the intervention of siRNA-OX40L, OX40L mRNA and protein levels were significantly inhibited (p <0.05), but the Foxp3 level was significantly increased in the experimental group (p <0.05). The IL-5 and TNF-α levels were significantly lower in the experimental group (p <0.05), and the reduction was more evident for the Th2-type cytokine IL-5 than for the Th1-type cytokine TNF-α. Few or no EOSs were found in the nasal mucosal epithelium of the experimental group (p <0.05), whereas EOS infiltration was significant in the other two groups. Blockage of the OX40/OX40L signaling pathway with siRNA-OX40L interference can inhibit allergic reactions and relieve allergic symptoms in AR mice. The underlying mechanism may be related to correcting Th2 immune deviation, inducing immune tolerance, and promoting Treg production. [ABSTRACT FROM AUTHOR]

This study determined the roles of OX40 and OX40L in women with recurrent spontaneous abortion (RSA). We compared the expression of OX40 and OX40L genes in peripheral blood mRNA levels and serum levels of OX40L in women with a history of RSA to the control group. In this case-control study, 40 women with a history of RSA (case group), and 40 others with no history of abortion (control group) were investigated. The expressions of OX40 mRNA and OX40L mRNA were determined in the two groups using the quantitative polymerase chain reaction. Also, the enzyme-linked immunosorbent assay was used to determine the levels of serum OX40L in the two groups. There were no significant differences in the maternal age of women in the two groups (30.1 ± 4.28 years in the case vs. 30.03 ± 4.23 years in the control group). There was no difference in terms of the levels of OX40 and OX40L mRNA between the groups (p = 0.08 and p = 0.56, respectively). In addition, there was no significant correlation between the expression of OX40 and OX40L mRNA levels with age or the number of abortions. The correlation between OX40 and OX40L mRNA levels was not significant. RSA history group turned to show a higher level of serum OX40L than the control group (p = 0.03). In conclusion, our findings demonstrated that the expression of OX40 mRNA and OX40L mRNA was similar between women with a history of RSA and the control group. The elevation of serum OX40L level may be considered as a risk factor for RSA. [ABSTRACT FROM AUTHOR]

Purpose: Epithelial thymic stromal lymphopoietin (TSLP) promotes Th2 inflammatory responses through induction of OX40 ligand (OX40L) on dendritic cells in allergic rhinitis (AR). Emerging evidence supports the important role of histamine H4 receptor (H4R) in allergic inflammation. This study aimed to investigate the effects of H4R in Th2-cytokine profile mediated by TSLP in AR. Methods: Human nasal epithelial cells (HNECs) from AR patients were stimulated with histamine in the presence or absence of H4R agonist (4-methylhistamine, 4-MH) and antagonist (NJ7777120, JNJ) or H1R agonist (2-pyridylethylamine). TSLP protein was measured by Western blotting and ELISA. To further elucidate the role of H4R in the in vivo situation of experimental AR, rats were sensitized and treated with JNJ or 4-MH. TSLP and OX40 ligand (OX40L) in the nasal mucosa were assayed by Western blotting. Th2 cytokines including interleukin-4, 5 and 13 in nasal lavage fluids were detected by ELISA. Results: Histamine alone did not induce TSLP production by HNECs. The pre-incubation with 4-MH prior to histamine promoted TSLP expression, which was inhibited by the stimulation with JNJ prior to histamine and 4-MH. The pre-incubation with 2-pyridylethylamine before histamine stimulation had no impact on TSLP production. In AR rats, the levels of TSLP and OX40L protein were increased as well as Th2 cytokines, which was further up-regulated by 4-MH treatment, while JNJ treatment attenuated these effects. Conclusions: H4R activation induced TSLP production by HNECs, which up-regulated OX40L expression in the nasal mucosa of sensitized rats. These factors promoted Th2-cytokine profile in AR. [ABSTRACT FROM AUTHOR]

Background The functional relevance of OX40 ligand (OX40L) in the effector phase of crescentic glomerulonephritis (GN) is unknown. These studies defined the role of endogenous OX40L during the effector stage of murine crescentic GN. Methods GN was induced by immunization with sheep globulin/adjuvant on Day 0 and injection of sheep anti-mouse glomerular basement membrane immunoglobulin (Ig) on Day 10. Rat IgG or neutralizing anti-OX40L antibody was administered on Days 10–18 and immune responses and renal injury assessed on Day 20. Results Compared with naïve animals, OX40L was upregulated in the lymph nodes (LNs) and on leucocytes and resident non-immune cells in the kidneys of mice with GN. Inhibition of OX40L in GN augmented renal injury, as indicated by increased crescent formation, proteinuria and glomerular leucocyte accumulation. In line with increased injury, anti-OX40L treatment increased proliferation and decreased apoptosis of CD4 T cells in the LNs, without affecting LN CD4 cytokine production and CD8 T-cell responses. Blockade of OX40L decreased LN regulatory T-cell (Treg) proliferation, transforming growth factor β production and foxp3 expression. OX40L inhibition did not affect B cell expansion or circulating antibody levels. In the kidney, neutralization of OX40L augmented interferon γ (IFNγ) expression by CD4 and CD8 T cells and shifted macrophage polarization towards the pro-inflammatory M1 phenotype. Conclusions OX40L is protective during the effector phase of murine crescentic GN by reducing the expansion of CD4 T cells and enhancing Treg responses in the LNs, and by locally inhibiting T-cell IFNγ production and pro-inflammatory macrophage phenotype in the kidney. [ABSTRACT FROM AUTHOR]

Abstract Influenza virus epidemics potentially cause pneumonia, which is responsible for much of the mortality due to the excessive immune responses. The role of costimulatory OX40–OX40 ligand (OX40L) interactions has been explored in the non‐infectious pathology of influenza pneumonia. Here, we describe a critical contribution of OX40L to infectious pathology, with OX40L deficiency, but not OX40 deficiency, resulting in decreased susceptibility to influenza viral infection. Upon infection, bronchiolar progenitors increase in number for repairing the influenza‐damaged epithelia. The OX40L expression is induced on the progenitors for the antiviral immunity during the infectious process. However, these defense‐like host responses lead to more extensive infection owing to the induced OX40L with α‐2,6 sialic acid modification, which augments the interaction with the viral hemagglutinin. In fact, the specific antibody against the sialylated site of OX40L exhibited therapeutic potency in mitigating the OX40L‐mediated susceptibility to influenza. Our data illustrate that the influenza‐induced expression of OX40L on bronchiolar progenitors has pathogenic value to develop a novel therapeutic approach against influenza.

Asthma, characterized by persistent inflammation and increased sensitivity of the airway, is the most common chronic condition among children. Novel, safe, and reliable treatment strategies are the focus of current research on pediatric asthma. Amygdalin, mainly present in bitter almonds, has anti-inflammatory and immunoregulatory potential, but its effect on asthma remains uninvestigated. Here, the impact of amygdalin on the thymic stromal lymphopoietin (TSLP)-dendritic cell (DC)-OX40L axis was investigated. A BALB/c mouse model for allergic asthma was established using the ovalbumin-sensitization method. Amygdalin treatment was administered between days 21 and 27 of the protocol. Cell numbers and hematoxylin and eosin (H&E) staining in bronchoalveolar lavage fluid (BALF) were used to observe the impact of amygdalin on airway inflammation. TSLP, IL-4, IL-5, IL-13, and IFN-γ concentrations were determined via Enzyme-linked immunosorbent assay (ELISA). TSLP, GATA-3, and T-bet proteins were measured using western blotting. Cell-surface receptor expression on DCs (MHC II, CD80, and CD86) was assessed via flow cytometry. OX40L mRNA and protein levels were detected using western blotting and qRT-PCR, respectively. Amygdalin treatment attenuated airway inflammation decreased BALF TSLP levels, inhibited DC maturation, restrained TSLP-induced DC surface marker expression (MHCII, CD80, and CD86), and further decreased OX40L levels in activated DCs. This occurred together with decreased Th2 cytokine levels (IL-4, IL-5, and IL-13) and GATA3 expression, whereas Th1 cytokine (IFN-γ) levels and T-bet expression increased. Amygdalin thus regulates the Th1/Th2 balance through the TSLP-DC-OX40L axis to participate in inflammation development in the airways, providing a basis for potential allergic asthma treatments.

A member of the short-chain dehydrogenase/reductase superfamily (DHRS1, SDR19C1) is a member of the short-chain dehydrogenase/reductase superfamily and a potential predictor of hepatocellular carcinoma (HCC). However, the role of DHRS1 in HCC immunity remains unclear. We systematically analyzed the association between DHRS1 and HCC immunity with transcriptional and clinical data from the Tumor Immune Estimation Resource, an integrated repository portal for tumor immune system interactions, and cBioPortal databases. Six DHRS1-associated immunomodulators strongly correlated with survival and were uncovered by exploiting univariate and multivariate Cox analyses. We created a risk score for each patient by adding the points from each immunomodulator and then classified them into high and low risk categories. Survival analysis were used to compare the overall survival between the 2 groups, and the receiver operating characteristic curve was applied to assess the accuracy of the risk score. Data from our center were adopted as the external validation set, the risk score was calculated using the risk coefficient of the 6 genes in the training cohort, and survival analysis were executed to verify the experimental group results. A nomogram was ultimately constructed with the R package. Our data revealed a correlation between the levels of immune cell infiltration and either the DHRS1 gene copy numbers or mRNA levels in HCC. Second, we generated a signature based on the 6 DHRS1-related immunomodulators (KDR, TNFRSF4, CD276, TNFSF4, SLAMF6, and SIGLEC9). We postulate that the generated risk scores would serve as an independent indicator of HCC prognosis, with an area under the receiver operating characteristic curve for the risk score of 0.743. We further established external validation sets to reconfirm the predictive validity of the risk score. Finally, a prognostic nomogram and calibration curve were created. The DHRS1 gene may exert an impact on HCC immunity. We posit that the nominated immune signature based on DHRS1-associated immunomodulators could constitute a promising prognostic biomarker in HCC., Competing Interests: The authors have no conflicts of interest to disclose., (Copyright © 2023 the Author(s). Published by Wolters Kluwer Health, Inc.)

Objective: To investigate the ferroptosis-related long non-coding RNAs (FRLncs) implicated in influencing the prognostic and immune microenvironment in osteosarcoma (OS), and to establish a foundational framework for informing clinical decision making pertaining to OS management., Methods: Transcriptome data and clinical data pertaining to 86 cases of OS, the GSE19276, GSE16088 and GSE33382 datasets, and a list of ferroptosis-related genes (FRGs) were used to establish a risk prognostic model through comprehensive analysis. The identification of OS-related differentially expressed FRGs was achieved through an integrated analysis encompassing the aforementioned 86 OS transcriptome data and the GSE19276, GSE16088 and GSE33382 datasets. Concurrently, OS-related FRLncs were ascertained via co-expression analysis. To establish a risk prognostic model for OS, Univariate Cox regression analysis and Lasso Cox regression analysis were employed. Subsequently, a comprehensive evaluation was conducted, comprising risk curve analysis, survival analysis, receiver operating characteristic curve analysis and independent prognosis analysis. Model validation with distinct clinical subgroups was performed to assess the applicability of the risk prognostic model to diverse patient categories. Moreover, single sample gene set enrichment analysis (ssGSEA) was conducted to investigate variations in immune cell populations and immune functions within the context of the risk prognostic model. Furthermore, an analysis of immune checkpoint differentials yielded insights into immune checkpoint-related genes linked to OS prognosis. Finally, the risk prognosis model was verified by dividing the samples into train group and test group., Results: We identified a set of seven FRLncs that exhibit potential as prognostic markers and influence factors of the immune microenvironment in the context of OS. This ensemble encompasses three high-risk FRLncs, denoted as APTR, AC105914.2 and AL139246.5, alongside four low-risk FRLncs, designated as DSCR8, LOH12CR2, AC027307.2 and AC025048.2. Furthermore, our analysis revealed notable down-regulation in the high-risk group across four distinct immune cell types, namely neutrophils, natural killer cells, plasmacytoid dendritic cells and tumor-infiltrating lymphocytes. This down-regulation was also reflected in four key immune functions, antigen-presenting cell (APC)-co-stimulation, checkpoint, cytolytic activity and T cell co-inhibition. Additionally, we identified seven immune checkpoint-associated genes with significant implications for OS prognosis, including CD200R1, HAVCR2, LGALS9, CD27, LAIR1, LAG3 and TNFSF4., Conclusion: The findings of this study have identified FRLncs capable of influencing OS prognosis and immune microenvironment, as well as immune checkpoint-related genes that are linked to OS prognosis. These discoveries establish a substantive foundation for further investigations into OS survival and offer valuable insights for informing clinical decision making in this context., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Breast cancer, which is the most common malignant tumor among women worldwide and an important cause of death in women. The existing prognostic model for patients with breast cancer is not accurate as breast cancer is resistant to commonly used antitumor drugs. Ferroptosis is a novel mechanism of programmed cell death that depends on iron accumulation and lipid peroxidation. Various studies have confirmed the role of ferroptosis in tumor regulation and ferroptosis is now considered to play an important role in breast cancer development. At present, the association between breast cancer prognosis and ferroptosis-related gene expression remains unclear. Further exploration of this research area may optimize the evaluation and prediction of prognosis of patients with breast cancer and finding of new therapeutic targets. In this study, clinical factors and the expression of multiple genes were evaluated in breast cancer samples from the Cancer Genome Atlas (TCGA) database and Gene Expression Omnibus (GEO) database database. Eleven prognostication-related genes (TP63, IFNG, MT3, ANO6, FLT3, PTGS2, SLC1A4, JUN, SLC7A5, CHAC1, and TF) were identified from differentially expressed genes to construct a survival prediction model, which showed a good prediction ability. KEGG pathway analysis revealed that immune-related pathways were the primary pathways. ssGSEA analysis showed significant differences in the distribution of certain immune-related cell subsets, such as CD8 + T cells and B cells, and in the expression of multiple immune genes, including type II IFN response and APC coinhibition. In addition, 10 immune targets related to ferroptosis in breast cancer were found: CD276, CD80, HHLA2, LILRA2, NCR3LG1, NECTIN3, PVR, SLAMF9,TNFSF4, and BTN1A1. Using TCGA, new ferroptosis genes related to breast cancer prognosis were identified, a new reliable and accurate prognosis model was developed, and 10 new potential therapeutic targets different from the traditional targeted drugs were identified to provide a reference for improving the poor prognosis of patients with breast cancer., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Purpose: The purpose of this study was to identify a prognostic signature based on stemness-related differentially expressed lncRNAs in colorectal cancer (CRC) and to investigate their potential as biomarkers for diagnosis, prognosis, and therapeutic targets., Methods: Stemness-related genes were collected from the TCGA cohort, and 13 differently expressed stemness-related lncRNAs were identified as prognostic factors for CRC using Kaplan-Meier analysis. A risk model was constructed based on the calculated risk score as a novel independent prognostic factor for CRC patients. The study also investigated the association between the risk model and immune checkpoints and m6A differentiation gene expression. qRT-PCR analysis was performed to validate the expression of differentially expressed stemness-related lncRNAs in CRC cell lines compared to normal colon mucosal cell line., Results: The low-risk lncRNAs were associated with higher survival in CRC patients (Kaplan-Meier analysis, P < 0.001). The risk model was a significant independent prognostic factor for CRC patients. Type I INF response was statistically significant between low- and high-risk groups. CD44, CD70, PVR, TNFSF4, BTNL2, CD40, these immune checkpoints were expressed differently between two risk groups. There was a significant difference between m6A differentiation gene expression such as METTL3, METTL14, WTAP, RBM15, ZC3H13, YTHDC2, YTHDF2, ALKBH5. qRT-PCR analysis validated that there were five up-regulated and eight down-regulated differently expressed stemness-related lncRNAs in CRC cell lines compared to the normal colon mucosal cell line., Conclusion: This study suggests that the 13 CRC stemness-related lncRNA signature could become a promising and reliable prognostic factor for colorectal cancer. The risk model based on the calculated risk score may have implications for personalized medicine and targeted therapies for CRC patients. The study also suggests that immune checkpoints and m6A differentiation genes may play important roles in the development and progression of CRC., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The development and progression of chronic hepatitis are closely associated with the immune function of human body, and T cells play an important role in this process. Costimulatory molecules OX40/OX40L are involved in the activation of T cells. This article reviews the molecular features of OX40/OX40L, the mechanism of action of OX40/OX40L in immune regulation, and the association between OX40/OX40L and chronic liver diseases. [ABSTRACT FROM AUTHOR]

The aim of the present study was to explore the roles of OX40/OX40 ligand (OX40L) signaling and OX40+ T cells in ovalbumin (OVA)‑induced mouse asthma model. Asthma was induced by OVA exposure and subsequent co‑treatment with OX40L protein, neutralizing anti‑OX40L blocking antibody, OX40+ T cells or PBS. The protein expression levels of interleukin (IL)‑4, IL‑6, IL‑13, IL‑17, tumor necrosis factor (TNF)‑α and interferon (IFN)‑γ in bronchoalveolar lavage fluid (BALF) were examined using murine cytokine‑specific ELISA. Eosinophil accumulation as well as proliferation and apoptosis of T cells in BALF were detected by Cell Counting kit‑8 and flow cytometric assays. Expression of the apoptosis‑related protein cleaved caspase‑3 was examined in OX40+ T cells using western blot assay. Flow cytometric analysis revealed that OVA‑treated mice that were co‑treated with OX40L or OX40+ T cells exhibited higher eosinophil infiltration compared with control mice treated only with OVA, whereas neutralizing anti‑OX40L blocking antibody inhibited eosinophil infiltration. ELISA assays demonstrated that the expression of IL‑4, IL‑6, IL‑13, IL‑17, TNF‑α and IFN‑γ in BALF in OX40L‑treated and OX40+ T cell‑treated mice was increased compared with expression levels in control mice. Treatment with OX40L protein effectively reduced apoptosis of T cells and the expression of cleaved caspase‑3 in T cells. OX40L‑treated and OX40+ T cell‑treated mice exhibited increased asthma through OX40/OX40L signaling, which probably promoted inflammatory factor expression, eosinophil infiltration and T cell proliferation. [ABSTRACT FROM AUTHOR]